This invention relates generally to chemical functionalization of surfaces to modify the properties thereof. More particularly, the invention relates to functionalization of a substrate with a silane mixture to reduce surface energy and thus constrain droplets of liquid that are applied to the substrate surface. A primary use of the invention is in the field of solid phase chemical synthesis, particularly solid phase synthesis of oligomer arrays.
Chemically modified, xe2x80x9cfunctionalized,xe2x80x9d solid surfaces are necessary in many laboratory procedures involved in chemistry and biotechnology. One important application is in solid phase chemical synthesis, wherein initial derivatization of a substrate surface enables synthesis of polymers such as oligonucleotides and peptides on the substrate itself. Support-bound oligomer arrays, particularly oligonucleotide arrays, may be used in screening studies for determination of binding affinity and in diagnostic applications, i.e., to detect the presence of a nucleic acid containing a specific, known oligonucleotide sequence. Modification of surfaces for use in chemical synthesis has been described, for example, in U.S. Pat. No. 5,624,711 to Sundberg et al., in U.S. Pat. No. 5,266,222 to Willis et al., in U.S. Pat. No. 5,137,765 to Farnsworth, and in numerous other patents and publications.
In modifying siliceous or metal oxide surfaces, one technique that has been used is derivatization with bifunctional silanes, i.e., silanes having a first functional group enabling covalent binding to the surface (often an Si-halogen or Si-alkoxy group, as in xe2x80x94SiCl3 or xe2x80x94Si(OCH3)3, respectively) and a second functional group that can impart the desired chemical and/or physical modifications to the surface. A problem with this type of surface modification, however, is that incorporation of a desirable surface chemical functionalityxe2x80x94provided by the second functional groupxe2x80x94may result in a surface with undesirable physical properties. For example, there is currently a great deal of interest in synthesizing arrays of different oligonucleotides on siliceous surfaces, and a high density of array features is generally considered desirable. The various array features can be independently created by the planar separation of individual phosphoramidite coupling reactions as the oligonucleotides are synthesized; a simple way to achieve this separation is by spotting the phosphoramidite solutions onto the surface. Feature density is then determined by the spread of the solution droplet, which is in turn uniquely determined by both the volume of the droplet and the contact angle between the droplet and the surface. However, to covalently couple the first nucleotide phosphoramidite to the substrate surface requires hydroxyl moieties on the surface, which makes the surface wettable by the phosphoramidite solutions and thus creates droplet spread; for a given droplet volume, then, relatively large array features are provided, limiting feature density.
The aforementioned problem can be overcome using a variety of techniques to constrain the droplets as they are applied to the substrate surface. Permanent wells can be formed by micromachining the substrate, with the active surfaces subsequently modified, constraining the droplet by capillary action. Temporary wells can also be formed using either a pre-formed xe2x80x9cstencilxe2x80x9d or by applying a coating to the substrate and patterning the coating. These wells could constrain the droplet by either capillary action and/or by using a relatively unwettable coating. Alternatively, as described in U.S. Pat. No. 5,474,796 to Brennan, a pattern of two different surface-bound silanes can be formed by physically masking the surface, depositing the first silane, and then removing the mask and depositing the second silane. This procedure can be used to constrain a droplet by surrounding a reactive spot on the surface, formed by one of the two silanes, with a lower surface energy spot, formed by the other of the two silanes.
All of these procedures, however, require considerable processing and thus add substantially to the time and cost required to fabricate an array. Also, the existence of a pattern on the substrate requires that the array writing apparatus be aligned with the surface pattern, a non-trivial issue for small array features.
The present invention is directed to the aforementioned need in the art, and provides a way of functionalizing substrate surfaces to reduce surface energy and thus constrain droplets of liquid that are applied to the substrate surface, while avoiding the aforementioned problems and difficulties associated with the procedures of the prior art.
Accordingly, it is a primary object of the invention to address the aforementioned need in the art and provide a relatively simple, straightforward process for preparing a low surface energy functionalized surface on a substrate.
It is an additional object of the invention to provide such a process by coupling a mixture of silanes to hydrophilic moieties present on a substrate surface.
It is another object of the invention to provide a process for preparing support-bound cleavable ligands on a low surface energy substrate, wherein the ligands may be small molecules, oligonucleotides, oligopeptides, or the like.
It is another object of the invention to provide a derivatizing composition for preparing a low surface energy functionalized surface on a substrate.
It is still another object of the invention to provide such a derivatizing composition comprising a mixture of silanes.
It is yet another object of the invention to provide such a derivatizing composition comprising a first silane that upon binding to a substrate reduces the surface energy thereof, and a second silane that upon binding to a substrate provides a means for covalently binding molecular moieties to the substrate surface.
It is a further object of the invention to provide substrates having low surface energy functionalized surfaces.
Additional objects, advantages and novel features of the invention will be set forth in part in the description which follows, and in part will become apparent to those skilled in the art upon examination of the following, or may be learned by practice of the invention.
In one embodiment of the invention, then, a process is provided for preparing a low surface energy functionalized surface on a substrate, which comprises contacting a substrate having reactive hydrophilic moieties on its surface with a derivatizing composition comprising a first silane R1xe2x80x94Si(RLRxRy) and a second silane R2xe2x80x94(L)nxe2x80x94Si(RLRxRy) under reaction conditions effective to couple the silanes to the substrate surface and provide xe2x80x94Sixe2x80x94R1 groups and xe2x80x94Sixe2x80x94(L)nxe2x80x94R2 groups thereon. The RL, which may be the same or different, are leaving groups, the Rx and Ry, which may also be the same or different, are either leaving groups, like RL, or are lower alkyl, R1 is a chemically inert moiety that upon binding to the substrate surface lowers the surface energy thereof, n is 0 or 1, L is a linking group, and R2 comprises either a functional group enabling covalent binding of a molecular moiety or a group that may be modified to provide such a functional group. The ratio of the silanes in the derivatizing composition determines the surface energy of the functionalized substrate and the density of molecular moieties that can ultimately be bound to the substrate surface.
In another embodiment, a process is provided for preparing support-bound cleavable ligands on a low surface energy substrate. The process involves contacting a substrate having reactive hydrophilic moieties on the surface thereof with a derivatizing composition comprising a first silane R1xe2x80x94Si(RLRxRy) and a second silane R2xe2x80x94(L)nxe2x80x94Si(RLRxRy) as described above, under reaction conditions effective to couple the silanes to the substrate surface and provide xe2x80x94Sixe2x80x94R1 groups and xe2x80x94Sixe2x80x94(L)nxe2x80x94R2 groups thereon. A ligand is then coupled to the surface at R2, through a linking moiety containing a cleavable site. The ligand may be, for example, a small molecule, a first monomer in the solid phase synthesis of an oligomer, an intact oligomer, or the like.
In an additional embodiment, a derivatizing composition is provided for carrying out the aforementioned processes. The derivatizing composition comprises a mixture of silanes, including a first silane R1xe2x80x94Si(RLRxRy) and a second silane R2xe2x80x94(L)nxe2x80x94Si(RLRxRy), wherein R1, R2, RL, Rx, Ry and n are as defined above.
Finally, the functionalized substrates provided using the presently disclosed and claimed processes and compositions represent a further embodiment of the invention. The substrates have surface-bound xe2x80x94Sixe2x80x94R1 groups and xe2x80x94Sixe2x80x94(L)nxe2x80x94R2 groups, wherein the R1 moieties reduce surface energy and the R2 moieties comprise either functional groups enabling covalent attachment of a molecular moiety of interest or modifiable groups that can be converted to such functional groups.